High affinity association of myo-inositol trisphosphates with phytase and its effect upon the catalytic potential of the enzyme

A neutral phytase from germinating mung bean (Vigna radiata) seeds dephosphorylatesmyo-inositol hexakisphosphate sequentially tomyo-inositol. The enzyme also binds with higher affinity tomyo-inositol trisphosphates (1,4,5), (2,4,5), and (1,3,4) isomers without catalysis. The high affinity complex elicits Ca2+ mobilization in vitro from microsomes/vacuoles via the formation of a ternary complex with the receptor for Ins(1,4,5)P3. As a sequel to our previous report, we have carried out a detailed characterization of the two sites and examined the mutual interactions between them. Presaturation of the high affinity site leads to an increase in the affinity of the enzyme for phytic acid and its rate of dephosphorylation as well. From the products of limited tryptic cleavage of phytase, two peptides, each with one activity, have been isolated. The larger peptide (≈ 66 kDa) contains the catalytic site, and the smaller peptide (≈ 5 kDa) has the high affinity myo-inositol trisphosphate-binding site. The interaction between the dual activities of phytase has been observed also at the level of the two peptides. A sequence homology search using N-terminal 12 amino acid residues of the 5-kDa fragment has revealed significant homology with the Homer class of proteins implicated in signaling pathways involving metabotropic glutamate receptor and myo-inositol 1,4,5-trisphosphate receptor. These results indicate a second role of phytase in Ca2+mobilization during germination of mung been seed via a salvage pathway that involves allosteric activation by myo-inositol trisphosphate

Effect of DNA Groove Binder Distamycin A upon Chromatin Structure

BACKGROUND: Distamycin A is a prototype minor groove binder, which binds to B-form DNA, preferentially at A/T rich sites. Extensive work in the past few decades has characterized the binding at the level of double stranded DNA. However, effect of the same on physiological DNA, i.e. DNA complexed in chromatin, has not been well studied. Here we elucidate from a structural perspective, the interaction of distamycin with soluble chromatin, isolated from Sprague-Dawley rat. METHODOLOGY/PRINCIPAL FINDINGS: Chromatin is a hierarchical assemblage of DNA and protein. Therefore, in order to characterize the interaction of the same with distamycin, we have classified the system into various levels, according to the requirements of the method adopted, and the information to be obtained. Isothermal titration calorimetry has been employed to characterize the binding at the levels of chromatin, chromatosome and chromosomal DNA. Thermodynamic parameters obtained thereof, identify enthalpy as the driving force for the association, with comparable binding affinity and free energy for chromatin and chromosomal DNA. Reaction enthalpies at different temperatures were utilized to evaluate the change in specific heat capacity (ΔCp), which, in turn, indicated a possible binding associated structural change. Ligand induced structural alterations have been monitored by two complementary methods--dynamic light scattering, and transmission electron microscopy. They indicate compaction of chromatin. Using transmission electron microscopy, we have visualized the effect of distamycin upon chromatin architecture at di- and trinucleosome levels. Our results elucidate the simultaneous involvement of linker bending and internucleosomal angle contraction in compaction process induced by distamycin. CONCLUSIONS/SIGNIFICANCE: We summarize here, for the first time, the thermodynamic parameters for the interaction of distamycin with soluble chromatin, and elucidate its effect on chromatin architecture. The study provides insight into a ligand induced compaction phenomenon, and suggests new mechanisms of chromatin architectural alteration

Domestic Wheat Price Formation and Food Inflation in India

Inflation, especially in food prices, has been persistently high in India during the past twenty four months. This has been a source of concern to policy-makers. Fortunately, food price increases are now starting to ease, after the major spike that occurred in the wake of the severe drought of 2009. However, there still remains concern that we: (a) need to better understand the factors that drive such spikes in key prices; and (b) design more effective policies to prevent such future price spikes. The main approach to understanding inflation and its drivers has typically rested, on the whole, in assessing aggregate macroeconomic (aggregate supply and demand) conditions, which then typically leads to consideration of macroeconomic and monetary) policies as the principal tool to deal with inflation surges. That may indeed be appropriate in most circumstances, but is often a blunt, sometimes costly instrument that can stifle growth, especially if price pressures arise from (temporary) supply constraints. Therefore, it may be important to complement an aggregate macroeconomic analysis of inflation with microeconomic analysis: to ascertain if inflation is being driven by specific price spikes in important food and non-food commodities, which has the potential to drive other commodity prices in a cost-push manner. This paper, on global wheat market developments, price transmission and impacts on Indian domestic markets, as well as an assessment of public policies to manage domestic prices, is part of a larger effort to improve our in-house (Department of Economic Affairs) research---to track, monitor and forecast fast-moving key macro-economic variables with potentially large consequences for public policy. We have begun to intensify our efforts. We are investing further systematically---to understand growth and inflation dynamics in the context of rising food inflationary pressures in India and worldwide. We are capturing more high frequency data, and applying quantitative modeling tools (as evident in our current Economic Survey). We take up wheat in this paper, because of recent rapid price rises globally, as well as domestically, and because it constitutes a major element of the overall wholesale and consumer food price inflation indices. Some aspects of the price formation and policy intervention processes in wheat are also likely to be structurally similar for other similar classes of important food items (such as rice), permitting broader insights. Our paper draws upon existing theoretical insights and modeling attempts in the literature; it is, nevertheless, useful to note three “biases” in our approach: (a) favoring analysis of short-term, high-frequency price formation (daily, monthly, or quarterly), versus alternative longer-term annual, structural models; (b) favoring simplified reduced form forecasting models that track high-frequency turning points well, over more elaborate models and tests of longerduration time-series data (which may tend to be more historical and backward-looking, and less useful for short-term forecasting); and (c) assessing current India-specific public interventions in greater detail, than in more general academic papers and models

Domestic Wheat Price Formation and Food Inflation in India

Inflation, especially in food prices, has been persistently high in India during the past twenty four months. This has been a source of concern to policy-makers. Fortunately, food price increases are now starting to ease, after the major spike that occurred in the wake of the severe drought of 2009. However, there still remains concern that we: (a) need to better understand the factors that drive such spikes in key prices; and (b) design more effective policies to prevent such future price spikes. The main approach to understanding inflation and its drivers has typically rested, on the whole, in assessing aggregate macroeconomic (aggregate supply and demand) conditions, which then typically leads to consideration of macroeconomic and monetary) policies as the principal tool to deal with inflation surges. That may indeed be appropriate in most circumstances, but is often a blunt, sometimes costly instrument that can stifle growth, especially if price pressures arise from (temporary) supply constraints. Therefore, it may be important to complement an aggregate macroeconomic analysis of inflation with microeconomic analysis: to ascertain if inflation is being driven by specific price spikes in important food and non-food commodities, which has the potential to drive other commodity prices in a cost-push manner. This paper, on global wheat market developments, price transmission and impacts on Indian domestic markets, as well as an assessment of public policies to manage domestic prices, is part of a larger effort to improve our in-house (Department of Economic Affairs) research---to track, monitor and forecast fast-moving key macro-economic variables with potentially large consequences for public policy. We have begun to intensify our efforts. We are investing further systematically---to understand growth and inflation dynamics in the context of rising food inflationary pressures in India and worldwide. We are capturing more high frequency data, and applying quantitative modeling tools (as evident in our current Economic Survey). We take up wheat in this paper, because of recent rapid price rises globally, as well as domestically, and because it constitutes a major element of the overall wholesale and consumer food price inflation indices. Some aspects of the price formation and policy intervention processes in wheat are also likely to be structurally similar for other similar classes of important food items (such as rice), permitting broader insights. Our paper draws upon existing theoretical insights and modeling attempts in the literature; it is, nevertheless, useful to note three “biases” in our approach: (a) favoring analysis of short-term, high-frequency price formation (daily, monthly, or quarterly), versus alternative longer-term annual, structural models; (b) favoring simplified reduced form forecasting models that track high-frequency turning points well, over more elaborate models and tests of longerduration time-series data (which may tend to be more historical and backward-looking, and less useful for short-term forecasting); and (c) assessing current India-specific public interventions in greater detail, than in more general academic papers and models

Global gene expression profiling data analysis reveals key gene families and biological processes inhibited by Mithramycin in sarcoma cell lines

AbstractThe role of Mithramycin as an anticancer drug has been well studied. Sarcoma is a type of cancer arising from cells of mesenchymal origin. Though incidence of sarcoma is not of significant percentage, it becomes vital to understand the role of Mithramycin in controlling tumor progression of sarcoma. In this article, we have analyzed the global gene expression profile changes induced by Mithramycin in two different sarcoma lines from whole genome gene expression profiling microarray data. We have found that the primary mode of action of Mithramycin is by global repression of key cellular processes and gene families like phosphoproteins, kinases, alternative splicing, regulation of transcription, DNA binding, regulation of histone acetylation, negative regulation of gene expression, chromosome organization or chromatin assembly and cytoskeleton

Naphthoquinone-mediated inhibition of lysine acetyltransferase KAT3B/p300, basis for non-toxic inhibitor synthesis

Hydroxynaphthoquinone-based inhibitors of the lysine acetyltransferase KAT3B (p300), such as plumbagin, are relatively toxic. Here, we report that free thiol reactivity and redox cycling properties greatly contribute to the toxicity of plumbagin. A reactive 3rd position in the naphthoquinone derivatives is essential for thiol reactivity and enhances redox cycling. Using this clue, we synthesized PTK1, harboring a methyl substitution at the 3rd position of plumbagin. This molecule loses its thiol reactivity completely and its redox cycling ability to a lesser extent. Mechanistically, non-competitive, reversible binding of the inhibitor to the lysine acetyltransferase (KAT) domain of p300 is largely responsible for the acetyltransferase inhibition. Remarkably, the modified inhibitor PTK1 was a nearly non-toxic inhibitor of p300. The present report elucidates the mechanism of acetyltransferase activity inhibition by 1,4-naphthoquinones, which involves redox cycling and nucleophilic adduct formation, and it suggests possible routes of synthesis of the non-toxic inhibitor

In vivo experiments demonstrate the potent antileishmanial efficacy of repurposed suramin in visceral leishmaniasis

Background: Treatment failure and resistance to the commonly used drugs remains a major obstacle for successful chemotherapy against visceral leishmaniasis (VL). Since the development of novel therapeutics involves exorbitant costs, the effectiveness of the currently available antitrypanosomatid drug suramin has been investigated as an antileishmanial, specifically for VL,in vitro and in animal model experiments. Methodology/Principal: Leishmania donovani promastigotes were treated with suramin and studies were performed to determine the extent and mode of cell mortality, cell cycle arrest and other in vitro parameters. In addition, L. donovani infected BALB/c mice were administered suramin and a host of immunological parameters determined to estimate the antileishmanial potency of the drug. Finally, isothermal titration calorimetry (ITC) and enzymatic assays were used to probe the interaction of the drug with one of its putative targets namely parasitic phosphoglycerate kinase (LmPGK). Findings: The in vitro studies revealed the potential efficacy of suramin against the Leishmania parasite. This observation was further substantiated in the in vivo murine model, which demonstrated that upon suramin administration, the Leishmania infected BALB/c mice were able to reduce the parasitic burden and also generate the host protective immunological responses. ITC and enzyme assays confirmed the binding and consequent inhibition of LmPGK due to the drug. Conclusions/Significance: All experiments affirmed the efficacy of suramin against L. donovani infection, which could possibly lead to its inclusion in the repertoire of drugs against VL